The cancer suppressor protein p53 is known to be stabilized and activated by diverse cellular stresses such as heat shock, hypoxia, osmotic shock, and DNA damage, leading to the inhibition of cell growth and apoptosis (Ko and Prives, Genes Dev. 10: 1054-1072, 1996; Levine, Cell 88: 323-331, 1997; Oren, Cancer Biol. 5: 221-227, 1994). However, almost nothing is known as to how p53 selects the pathway of cell growth inhibition or apoptosis.
It has been also known that appptosis and cell cycle arrest are the major tumor suppressing function of p53 (Levine, Cell 88: 323-331, 1997). Therefore, the elucidation of mechanism of p53-induced apoptosis may contribute to the development of anticancer agents having a novel mechanism of action in the cancer treatment. Induction of apoptosis due to the expression of exogenous p53 has been reported in some, though not all, cancer cells with the p53 mutation (Gomez-Manzano etal., Cancer Res. 56: 694-699; Kock et al. , Int. J. Cancer 67: 808-815, 1996). Therefore, if the apoptosis-associated target gene of p53 is identified and allowed to express in cancer cells, or a protein encoded by the gene is introduced into the cancer cell, thereby inducing apoptosis, effective cancer treatment would be possible. Besides cancer, examples of apoptosis-associated disorders include arteriosclerosis, Alzheimer's disease, amyotrophic lateral sclerosis, graft-versus-host disease, autoimmune lymphocytosis syndrome, and viral infection. The identification of apoptosis-associated p53 target gene would greatly contribute to the elucidation of causes for these disorders or development of their effective therapy.
Furthermore, a mitochondrial protein bcl-2 has been known to inhibit the apoptosis process, promoting cell survival (Vaux, D. L. et al. , Nature, 335: 440-442, 1988; Tsujimoto, Y. Oncogene, 4: 1331-1336, 1989; Sentman, C. L. et al., Cell, 67: 879-888, 1991; Strasser, A. Cell, 67: 889-899, 1991). Bcl-2 gene was first isolated as an oncogene activated by the chromosome translocation in a human follicular lymphoma (Tsujimoto, Y. et al., Science (Washington D.C.), 226: 1097-1099, 1984; Bakhshi, A. Cell, 41: 899-906, 1985;Cleary, M. L. et al. , Proc. Natl. Acad. Sci. USA, 82:7439-7443, 1985). In Caenorhabditis elegans, ced-3 and ced-4 are essential for apoptosis during development, while ced-9 inhibits their actions (Ellis, H. M. and Horvitz, H. R. Cell, 44: 817-829, 1986; Hengartner, M. O. and Horvitz, H. R. Curr. Opin. Genet. Dev., 4: 581-586, 1994). Since bcl-2 is functionally as well as structurally a human homolog of ced-9 (Hengartner, M. O. and Horvitz, H. R. Cell, 76: 665-676, 1994), this mechanism of apoptosis seems to be extremely well conserved from nematodes to mammals. Although the major action mechanism of bcl-2 is uncertain, the bcl-2 gene product is thought to directly or indirectly interfere with the release of cytochrome c from mitochondria (Yang, J. et al., Science (Washington D.C.) 275: 1129-1132, 1997; Kluck, R. M. et al., Science (Washington D.C.), 275: 1132-1136, 1997; Shimizu, S., et al. , Nature, 399: 483-487, 1999). Bcl-2 contains four functional domains called BH1, BH2, BH3, and BH4 (Reed, J. C. Oncogene, 17: 3225-3236, 1998). More than seventeen distinct members of the bcl-2 family proteins have been reported as an anti-apoptosis factor or apoptosis inducer, all of which contain at least one of these four different domains.
The majority of mechanisms of the p53-dependent apoptosis, which is thought to be the most important characteristic of cancer suppression by p53, have not been elucidated. Several target genes have been isolated as those encoding candidate proteins associated with the p53-dependent apoptosis, including Bax (Miyashita, T. and Reed, J. C. Cell, 80: 293-299, 1995), PIG3 (Polyak, K., et al. , Nature, 389: 300-305, 1997; Venot, C. et al. , EMBO J. , 17: 4668-4679, 1998), Killer/DR5 (Wu, G. S., et al., Nat. Genet., 17: 141-143, 1997), Fas (Owen-Schaub, L. B., et al., Mol. Cell Biol., 15: 3032-3040, 1995; Muller, M., et al., J. Exp. Med., 188: 2033-2045, 1998), Noxa (Oda, E. , et al. , Science (Washington D.C.), 288: 1053-1058, 2000), PERP (Attardi, L. D., et al., Genes Dev., 14: 704-718, 2000), and PUMA (Nakano, K. and Vousden, K. H., Mol. Cell, 7: 683-694, 2001; Yu, J., et al., Mol. Cell, 7: 673-682, 2001). Bax, Noxa, and PUMA are mitochondrial proteins, containing the BH-3 domain, and belong to the bcl-2 family. PIG3 is a homolog of the plant NADPH oxidoreductase TED2 involved in the apoptosis process necessary for the formation of plant meristem. Killer/DR5 and Fas are receptors to transmit the external cell death signal. PERP is a cellular plasma membrane protein whose overproduction induces apoptosis. Indeed these known target genes encode candidate proteins involved in the p53-dependent apoptosis but they are not sufficient for explaining the mechanism of p53-induced apoptosis.
Knowledge obtained by elucidating the cancer suppression mechanism of p53, and in particular the mechanism of p53-mediated apoptosis, are expected to lead to the development of novel drugs having the different mechanism of action from the conventional one, especially apoptosis-mediated cancer therapeutic agents.